Inorganic phosphate plays a key role in a myriad of biological processes, including bone mineralization, reversible regulation of protein function by phosphorylation, and production of adenosine triphosphate. Plasma levels of phosphate range between 2.2 and 4.9 mg/dl (Dwyer et al., “Severe Hypophosphatemia in Postoperative Patients,” Nutr Clin Pract 7(6):279-283 (1992), Alon et al., “Calcimimetics as an Adjuvant Treatment for Familial Hypophosphatemic Rickets,” Clin J Am Soc Nephrol 3: 658-664 (2008)), and are primarily regulated by modifying renal tubular reabsorption. Because of phosphate's pleiotropic activity, imbalances in phosphate homeostasis adversely affect essentially every major tissue/organ.
Hypophosphatemia is a common clinical condition with an incidence ranging from 0.2-3.1% in all hospital admissions to 21.5-80% in specific subgroups of hospitalized patients (Gaasbeek et al., “Hypophosphatemia: An Update on its Etiology and Treatment,” Am J Med 118(10):1094-1101 (2005), Brunelli et al., “Hypophosphatemia: Clinical Consequences and Management,” J Am Soc Nephrol 18(7):1999-2003 (2007)). Acute clinical manifestations of hypophosphatemia include respiratory failure, cardiac arrhythmia, hemolysis, rhabdomyolysis, seizures, and coma. Chronic clinical manifestations of hypophosphatemia include myalgia and osteomalacia (Gaasbeek et al., “Hypophosphatemia: An Update on its Etiology and Treatment,” Am J Med 118(10):1094-1101 (2005)). Hypophosphatemia originates from diverse pathophysiologic mechanisms, most importantly from renal phosphate wasting, an inherited or acquired condition in which renal tubular reabsorption of phosphate is impaired (Imel et al., “Fibroblast Growth Factor 23: Roles in Health and Disease,” J Am Soc Nephrol 16(9):2565-2575 (2005); Negri A., “Hereditary Hypophosphatemias: New Genes in the Bone-kidney Axis,” Nephrology (Carlton) 12(4):317-320 (2007)). Hypophosphatemia can also be associated with alcoholic and diabetic ketoacidosis, acute asthma, chronic obstructive pulmonary disease, sepsis, recovery from organ transplantation, and the “refeeding syndrome”, which refers to metabolic disturbances seen in malnourished patients on commencing nutrition (Gaasbeek et al., “Hypophosphatemia: An Update on its Etiology and Treatment,” Am J Med 118(10):1094-1101 (2005), Miller et al., “Hypophosphatemia in the Emergency Department Therapeutics,” Am J Emerg Med 18(4):457-461 (2000), Marinella M A., “Refeeding Syndrome and Hypophosphatemia,” J Intensive Care Med 20(3):155-159 (2005)).
Oral or intravenous administration of inorganic phosphate salts is the current mainstay for the management of hypophosphatemia. Oral phosphate therapy requires high doses, which frequently lead to diarrhea or gastric irritation (Shiber et al., “Serum Phosphate Abnormalities in the Emergency Department,” J Emerg Med 23(4):395-400 (2002)). For intravenous phosphate therapy, the response to any given dose is sometimes unpredictable (Bohannon N J., “Large Phosphate Shifts with Treatment for Hyperglycemia,” Arch Intern Med 149(6):1423-1425 (1989), Charron et al., “Intravenous Phosphate in the Intensive Care Unit: More Aggressive Repletion Regimens for Moderate and Severe Hypophosphatemia,” Intensive Care Med 29(8):1273-1278 (2003); Rosen et al., “Intravenous Phosphate Repletion Regimen for Critically III patients with Moderate Hypophosphatemia,” Crit Care Med 23(7):1204-1210 (1995)), and complications include “overshoot” hyperphosphatemia, hypocalcemia, and metastatic calcification (Gaasbeek et al., “Hypophosphatemia: An Update on its Etiology and Treatment,” Am J Med 118(10):1094-1101 (2005); Shiber et al., “Serum Phosphate Abnormalities in the Emergency Department,” J Emerg Med 23(4):395-400 (2002)). In addition, parenteral regimens are not practical for chronic disorders. Most importantly, replacement therapy alone is never adequate when there is significant renal phosphate wasting. Therefore, novel strategies for the treatment of hypophosphatemia are needed.
Kidney transplantation is the preferred treatment of end-stage renal failure, and hypophosphatemia is a well recognized problem during the first weeks after engraftment. The majority of kidney transplant patients often experience excessive renal phosphate leakage (Schwarz et al., “Impaired Phosphate Handling of Renal Allografts is Aggravated under Rapamycin-based Immunosuppression,” Nephrol Dial Transplant 16: 378-382 (2001); Moorhead et al., “Hypophosphataemic Osteomalacia after Cadaveric Renal Transplantation,” Lancet 1(7860):694-697 (1974)), because the transplanted kidneys only marginally reabsorb the urinary phosphate to the circulation. The reasons for this poor reabsorbing activity on the part of transplanted kidneys are unknown. It frequently causes the patients malnutrition and secondary osteoporosis. This problem cannot be treated by a simple exogenous supplementation of phosphate. Similar renal phosphate leakage with unknown pathology is often observed in pediatric medicine, with outcomes such as malnutrition or growth retardation.
A recent study in adults demonstrated that as many as 93% of patients develop moderate to severe hypophosphatemia (serum phosphate concentration 0.9-2.25 mg/dL), an average of 5 weeks following transplantation (Ambuhl et al., “Metabolic Aspects of Phosphate Replacement Therapy for Hypophosphatemia After Renal Transplantation: Impact on Muscular Phosphate Content, Mineral Metabolism, and Acid/base Homeostasis,” Am J Kidney Dis 34:875-83 (1999)).
Health problems associated with circulating phosphate shortage are not limited to humans. Dairy cows sometimes suffer from hypophosphatemia (too low phosphate in the blood) caused by overproduction of the milk. It not only deteriorates the nutritional quality of the milk but also often make the cows useless for milk production. It is a relatively common problem in dairy farms (Goff, J P., “Pathophysiology of Calcium and Phosphorus Disorders,” Vet Clin North Am Food Anim Pract 16(2):319-37 (2000), Oetzel, G R., “Management of Dry Cows for the Prevention of Milk Fever and Other Mineral Disorders,” Vet Clin North Am Food Anim Pract 16(2):369-86 (2000)).
Fibroblast growth factor (FGF) 23, is an endocrine regulator of phosphate homeostasis, and was originally identified as the mutated gene in patients with the phosphate wasting disorder “autosomal dominant hypophosphatemic rickets” (ADHR) (Anonymous., “Autosomal Dominant Hypophosphataemic Rickets is Associated with Mutations in FGF23,” Nat Genet 26(3):345-348 (2000)). FGF23 inhibits reabsorption of phosphate in the renal proximal tubule by decreasing the abundance of the type II sodium-dependent phosphate transporters NaPi-2A and NaPi-2C in the apical brush border membrane (Baum et al., “Effect of Fibroblast Growth Factor-23 on Phosphate Transport in Proximal Tubules,” Kidney Int 68(3):1148-1153 (2005); Perwad et al., “Fibroblast Growth Factor 23 Impairs Phosphorus and Vitamin D Metabolism In Vivo and Suppresses 25-hydroxyvitamin D-1alpha-hydroxylase Expression In Vitro,” Am J Physiol Renal Physiol 293(5):F1577-1583 (2007); Larsson et al., “Transgenic mice expressing fibroblast growth factor 23 under the control of the alpha1(I) collagen promoter exhibit growth retardation, osteomalacia, and disturbed phosphate homeostasis,” Endocrinology 145(7):3087-3094 (2004)). The phosphaturic activity of FGF23 is down-regulated by proteolytic cleavage at the 176RXXR179 (SEQ ID NO: 1) motif, where “XX” is defined as “HT”, corresponding to positions 177 and 178, respectively, of the FGF23 amino acid sequence, producing an inactive N-terminal fragment (Y25 to R179) and a C-terminal fragment (S180 to I251) (FIG. 1A) (Goetz et al., “Molecular Insights into the Klotho-dependent, Endocrine Mode of Action of Fibroblast Growth Factor 19 Subfamily Members,” Mol Cell Biol 27(9):3417-3428 (2007)). FGF receptor (FGFR) 1 is the principal mediator of the phosphaturic action of FGF23 (Liu et al., “FGFR3 and FGFR4 do not Mediate Renal Effects of FGF23,” J Am Soc Nephrol 19(12):2342-2350 (2008); Gattineni et al., “FGF23 Decreases Renal NaPi-2a and NaPi-2c Expression and Induces Hypophosphatemia in vivo Predominantly via FGF Receptor 1,” Am J Physiol 297(2):F282-F291 (2009)). In addition, Klotho, a protein first described as an aging suppressor (Kuro-o et al., “Mutation of the Mouse Klotho Gene Leads to a Syndrome Resembling Aging,” Nature 390(6655):45-51 (1997)), is required as a coreceptor by FGF23 in its target tissue in order to exert its phosphaturic activity (Kurosu et al., “Regulation of Fibroblast Growth Factor-23 Signaling by Klotho,” J Biol Chem 281(10):6120-6123 (2006); Urakawa et al., “Klotho Converts Canonical FGF Receptor into a Specific Receptor for FGF23,” Nature 444(7120):770-774 (2006)). Klotho constitutively binds the cognate FGFRs of FGF23, and the binary FGFR-Klotho complexes exhibit enhanced binding affinity for FGF23 ((Kurosu et al., “Regulation of Fibroblast Growth Factor-23 Signaling by Klotho,” J Biol Chem 281(10):6120-6123 (2006); Urakawa et al., “Klotho Converts Canonical FGF Receptor into a Specific Receptor for FGF23,” Nature 444(7120):770-774 (2006)). In co-immunoprecipitation studies, it was demonstrated that the mature, full-length form of FGF23 (Y25 to I251) but not the inactive N-terminal fragment of proteolytic cleavage (Y25 to R179) binds to binary FGFR-Klotho complexes (Goetz et al., “Molecular Insights into the Klotho-dependent, Endocrine Mode of Action of Fibroblast Growth Factor 19 Subfamily Members,” Mol Cell Biol 27(9):3417-3428 (2007)).
The present invention is directed to overcoming the deficiencies in the art.